Cells are usually cultured in a bioreactor where the conditions may be controlled. Bioreactors may be a closed system so as to avoid contamination from the outside world. This is especially true for mammalian cells, and in particular for human cells. Control of nutrients, oxygen, temperature and pH are of vital importance for culturing cells.
In common used bioreactors there is both a liquid phase and a gas phase. The control of e.g. dissolved oxygen in the medium occurs at the interphase of the gas phase and liquid phase. Carbon dioxide may be used to control the pH. Some bioreactors have only a liquid phase, see e.g. WO2011/142667 and a device is needed to control dissolved oxygen and the pH. Transfer of gasses such as oxygen to a fluid medium may be performed over a membrane. Common oxygenators comprise of a chamber comprising oxygen where medium is flown through. EP0343357 discloses an oxygenator using 0.132 inch inner diameter (ID) and 0.183 inch outer diameter (OD) silicon tube where the fluid is flown through. Air is pumped by an air pump into the shell side of the oxygenator. Due to the gas permeable properties of silicone, oxygen is transferred to the fluid. The pH can be controlled by controlling the dissolved carbon dioxide in the medium. U.S. Pat. No. 4,493,692 discloses a blood gas concentration control apparatus for regulating oxygen an carbon dioxide in extracorporeal blood. It discloses an oxygenator wherein a source of oxygen (100% O2) and a source of carbon dioxide (95% O2 and 5% CO2) is connected. Both gasses are applied to the same oxygenator. WO2011/066481 discloses an extracorporeal membrane oxygenating control wherein an oxygen supply and carbon dioxide supply provide oxygen and carbon dioxide to the same membrane oxygenator.
The present inventors found that mixtures of oxygen and carbon dioxide and nitrogen, such as used in the prior art, give large fluctuations in the control of the pH and dissolved oxygen in a fluid. With a mixture of gasses, the pH could only be controlled within 0.5 pH units of the setpoint. Similarly, the control of the setpoint of the dissolved oxygen was in the range of 25% air saturation. For the culturing of mammalian cell, especially human cell and in particular human stem cells, these kind of controls are not tight enough to create a proper environment for the cells in a single phase bioreactor. In addition to the large variation of both the pH and dissolved oxygen around the setpoint, the control system for pH and dissolved oxygen was very slow.